Botulinum neurotoxin type A (BoNT/A), a potent paralytic, is a widely used treatment for the masseter muscles. Clinical reports suggest that some effects of BoNT/A treatment (atrophy, reduced electrical activity) are unusually long-lasting in jaw muscles, but the reasons for this are not clear. Possible explanations are reduction in the number of motor units or a poor response of the satellite cell population, but as yet no animal studies have been performed to study the biology of jaw muscles after toxin treatment. In clinical usage, bite force recovers more rapidly than the masseter, but the mechanism for this is unknown. Additionally, one benefit claimed for BoNT/A but unproven is that the jaw will be unloaded after muscle paralysis. If so, then disuse loss of bone should result, but this too is undemonstrated. The proposed research is a systematic examination of the effects of BoNT/A on the masseter muscle and the mandible, using two accepted animal models of human masticatory function, rabbit and pig. Specific Aim 1 will use electromyography and motion analysis to clarify whether compensatory patterns of muscle usage account for the early return of bite force and masticatory ability. Specific Aim 2 will assess the possible unloading of the injection side mandible by strain gage recording and microCT examination of bony architecture. Masseter paralysis is hypothesized to be followed rapidly by loss of bone mass in unloaded regions. Specific Aim 3 will investigate whether BoNT/A paralysis causes reorganization of motor units within the masseter and will employ neurotracing and nerve stimulation techniques. The immediate response to the toxin is expected to be expansion of normally highly restricted motor unit territories. The possibility of motor neuron loss will also be investigated. Specific Aim 4 will establish the course of muscle degeneration and repair to explore why atrophy is long-lasting in the masseter. The activity of the satellite cell population will be described through markers of replication and cell death and by tracking changes in myosin heavy chain composition. In summary, this project addresses the biology underlying a common clinical treatment. It will provide new information on how the muscles and their nerves are affected, how jaw biomechanics are altered, and the extent of neuroplasticity of the jaw motor system. The results will provide an informed background for patient treatment and for interpreting outcomes. PUBLIC HEALTH RELEVANCE: Botulinum neurotoxin type A, better known under the commercial name of Botox, paralyzes muscles for extended periods of time, during which the muscles atrophy and the nerves reorganize. Thousands of patients have received the drug in order to shrink large jaw muscles. The proposed research will use animal models to study how the chewing system adapts to the toxin and the biology of the regenerative response. The results will guide clinicians in understanding functional consequences and help to validate clinical rationales for treatment.